Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
  • C07H17/00—Compounds containing heterocyclic radicals directly attached to hetero atoms of saccharide radicals
  • C07H17/04—Heterocyclic radicals containing only oxygen as ring hetero atoms
  • C07H17/08—Hetero rings containing eight or more ring members, e.g. erythromycins
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/04—Antibacterial agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P33/00—Antiparasitic agents
  • A61P33/02—Antiprotozoals, e.g. for leishmaniasis, trichomoniasis, toxoplasmosis
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
  • C07H17/00—Compounds containing heterocyclic radicals directly attached to hetero atoms of saccharide radicals

Definitions

• This invention relates to novel macrolide compounds that are useful as antibacterial and antiprotozoal agents in mammals, including man, as well as in fish and birds.
• This invention also relates to methods of preparing the novel compounds, novel intermediates useful in preparation of the compounds, and pharmaceutical compositions containing the novel compounds.
• the present invention includes methods of treating bacterial and protozoal infections through the administration of the novel compounds to mammals, fish and birds requiring such treatment.
• the present invention relates to compounds of the formula and to pharmaceutically acceptable salts, prodrugs, tautomers, and solvates, thereof, wherein:
• Substituted groups in this list can be substituted with T substituents (i.e., with T substituents other than those indicated as “substituted”).
• R 1 is an alpha-branched C 3 -C 8 alkyl, alkenyl, alkynyl, alkoxyalkyl or alkylthioalkyl group any of which may optionally be substituted by one or more hydroxyl groups; C 5 -C 8 cycloalkylalkyl group wherein the alkyl group is an alpha-branched C 2 -C 5 alkyl group; C 3 -C 8 cycloalkyl or C 5 -C 8 cycloalkenyl group, either of which may optionally be substituted by methyl or one or more hydroxyl or one or more C 1 -C 4 alkyl groups or halo atoms; or a 3 to 6 membered oxygen or sulphur containing heterocyclic ring which may be saturated, or fully or partially unsaturated and which may optionally be substituted by one or more C 1 -C 4 alkyl groups or halo atoms; or R 1 is phenyl or C 6
• R 1 is Me, ethyl, n-butyl, MeS, EtS, cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.
• X 1 is -C(O)-.
• Ar is selected from phenyl, 2-methoxyphenyl, 4-methoxyphenyl, quinolin-4-yl, 7-methoxy-quinolin-4-yl, 4-phenyl-imidazol-1-yl, pyridin-4-yl, pyridin-3-yl, pyridin-2-yl, 4-pyridinyl-1H-imidazol-1-yl, imidazo(4,5-b)pyridin-3-yl, 2-phenyl-thiazol-5-yl, 2-pyridin-3-yl-thiazol-4-yl and benzimidazol-1-yl.
• B is selected from NH, NMe and CH 2
• R 2 is (CH 2 ) n Ar.
• n is 3.
• Ar is preferably selected from quinolin-4-yl, 4-phenyl-1H-imidazol-1-yl, imidazo(4,5-b)pyridin-3-yl and 4-pyridinyl-1H-imidazol-1-yl.
• R 1 is ethyl
• R 3 is H
• R 10 is -OC(O)NH 2
• X 1 is-C(O)-.
• B is (CR aa R bb ) m where m is 0 (i.e., B is a bond)
• R 2 is (CH 2 ) 4 -(5- to 10- membered heteroaryl).
• heteroaryl in said R 2 group is 4-pyridinyl-1H-imidazol-1-yl.
• R 1 is ethyl
• R 3 is H
• R 10 is -OC(O)NH 2
• X 1 is -C(O)-
• -B-R 2 is -NH(CH 2 ) 3 -(5- to 10- membered heteroaryl).
• Preferred compounds within this group are those wherein heteroaryl in said R 2 is quinolin-4-yl.
• R 2 is H
• B is (CR aa R bb ) m where m is 0 (i.e., B is a bond)
• R 1 ethyl is H
• R 3 is H
• X 1 is -C(O)-
• R 10 is OC(O)NHR 11 R 12 .
• the invention also relates to a pharmaceutical composition for the treatment of a bacterial infection or a protozoa infection in a mammal, fish, or bird which comprises a therapeutically effective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
• the invention also relates to a method of treating a bacterial infection or a protozoa infection in a mammal, fish, or bird which comprises administering to said mammal, fish or bird a therapeutically effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof.
• treatment includes the treatment or prevention of a bacterial infection or protozoa infection as provided in the method of the present invention.
• bacterial infection(s) and protozoa infection(s) include bacterial infections and protozoa infections that occur in mammals, fish and birds as well as disorders related to bacterial infections and protozoa infections that may be treated or prevented by administering antibiotics such as the compounds of the present invention.
• Such bacterial infections and protozoa infections, and disorders related to such infections include the following: pneumonia, otitis media, sinusitus, bronchitis, tonsillitis, and mastoiditis related to infection by Streptococcus pneumoniae, Haemophilus influenzae , Moraxella catarrhalis , Staphylococcus aureus, or Peptostreptococcus spp.; pharynigitis, rheumatic fever, and glomerulonephritis related to infection by Streptococcus pyogenes, Groups C and G streptococci, Clostridium diptheriae , or Actinobacillus haemolyticum; respiratory tract infections related to infection by Mycoplasma pneumoniae , Legionella pneumophila , Streptococcus pneumoniae , Haemophilus influenzae, or Chlamydia pneumoniae; uncomplicated skin and soft tissue infections, abscesses and osteomyelitis, and
• aureus food poisoning and Toxic shock syndrome
• Groups A, B, and C streptococci ulcers related to infection by Helicobacter pylori; systemic febrile syndromes related to infection by Borrelia recurrentis; Lyme disease related to infection by Borrelia burgdorferi; conjunctivitis, keratitis, and dacrocystitis related to infection by Chlamydia trachomatis, Neisseria gonorrhoeae , S . aureus, S . pneumoniae , S. pyogenes, H .
• MAC Mycobacterium avium complex
• gastroenteritis related to infection by Campylobacter jejuni
• intestinal protozoa related to infection by Cryptosporidium spp.
• odontogenic infection related to infection by viridans streptococci
• persistent cough related to infection by Bordetella pertussis
• gas gangrene related to infection by Clostridium perfringens or Bacteroides spp.
• atherosclerosis related to infection by Helicobacter pylori or Chlamydia pneumoniae.
• Bacterial infections and protozoa infections and disorders related to such infections that may be treated or prevented in animals include the following: bovine respiratory disease related to infection by P. haem ., P. multocida, Mycoplasma bovis, or Bordetella spp.; cow enteric disease related to infection by E . coli or protozoa (i.e., coccidia, cryptosporidia, etc.); dairy cow mastitis related to infection by Staph. aureus, Strep. uberis, Strep. agalactiae, Strep.
• dysgalactiae Klebsiella spp., Corynebacterium, or Enterococcus spp.
• swine respiratory disease related to infection by A. pleuro ., P. multocida , or Mycoplasma spp.
• swine enteric disease related to infection by E . coli , Lawsonia intracellularis , Salmonella, or Serpulina hyodyisinteriae
• cow footrot related to infection by Fusobacterium spp.
• cow metritis related to infection by E .
• the present invention also relates to a method of preparing the above compound of formula I, or a pharmaceutically acceptable salt thereof, which comprises treating a compound of the formula wherein X, R 1 , and R 2 are as defined above, and where P is a hydroxy protecting group, to remove the hydroxy protecting group.
• P is an acetyl group.
• the above compound of formula II is prepared by treating a compound of the formula with R 2 -B-NH 2 .
• Me refers to methyl
• hydroxy protecting group includes acetyl, benzyloxycarbonyl, and various hydroxy protecting groups familiar to those skilled in the art including the groups referred to in T. W. Greene, P. G. M. Wuts, "Protective Groups In Organic Synthesis,” (J. Wiley & Sons, 1991).
• halo as used herein, unless otherwise indicated, includes fluoro, chloro, bromo or iodo.
• alkyl as used herein, unless otherwise indicated, includes saturated monovalent hydrocarbon radicals having straight, cyclic or branched moieties, or mixtures thereof. It is to be understood that where cyclic moieties are intended, at least three carbons in said alkyl must be present. Such cyclic moieties include cyclopropyl, cyclobutyl and cyclopentyl.
• alkenyl includes straight-chain or branched-chain mono- or poly-unsaturated aliphatic hydrocarbon radicals containing at least one carbon-carbon double bond.
• alkenyl radicals include, but are not limited to, ethenyl, E- and Z-propenyl, isopropenyl, E- and Z-butenyl, E- and Z-isobutenyl, E- and Z-pentenyl, E- and Z-hexenyl, E,E-, E,Z-, Z,E- and Z,Z-hexadienyl and the like.
• alkynyl includes straight-chain or branched-chain mono- or poly-unsaturated aliphatic hydrocarbon radicals containing at least one carbon-carbon triple bond.
• alkynyl radicals include, but are not limited to, ethynyl, propynyl, isopropynyl, butynyl, isobutynyl, pentynyl, hexynyl and the like.
• alkoxy as used herein, unless otherwise indicated, includes -O-alkyl groups wherein alkyl is as defined above.
• aryl includes an organic radical derived from an aromatic hydrocarbon by the removal of one hydrogen.
• aryl radicals include, but are not limited to, phenyl, naphthyl, indenyl, indanyl, azulenyl, fluorenyl, anthracenyl and the like.
• 3- to 10-membered heterocyclic includes aromatic and non-aromatic heterocyclic groups containing one or more heteroatoms, each selected from O, S and N, wherein each heterocyclic group has from 3 to 10 atoms in its ring system.
• Non-aromatic heterocyclic groups include groups having only 3 atoms in their ring system, but aromatic heterocyclic groups must have at least 5 atoms in their ring system.
• the heterocyclic groups include benzo-fused ring systems and ring systems substituted with one or more oxygen or nitrogen atoms.
• the heterocyclic groups also include partially unsaturated or fully saturated 3- to 10-membered ring systems, e.g., single rings of 3 to 8 atoms in size and bi- or tricyclic ring systems, including aromatic 6-membered aryl or heteroaryl rings fused to a non-aromatic ring.
• An example of a 4-membered heterocyclic group is azetidinyl (derived from azetidine).
• An example of a 5-membered heterocyclic group is thiazolyl, and an example of a 10-membered heterocyclic group is quinolinyl.
• non-aromatic heterocyclic groups are pyrrolidinyl, tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, tetrahydrothiopyranyl, piperidino, morpholino, thiomorpholino, thioxanyl, piperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1,2,3,6-tetrahydropyridinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl, pyrazolinyl, dithianyl, dithiolanyl, dihydropyr
• aromatic heterocyclic groups are pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinox
• a group derived from pyrrole may be C-attached or N-attached where such is possible.
• a group derived from pyrrole may be pyrrol-1-yl (N-attached) or pyrrol-3-yl (C-attached).
• protecting group refers to a suitable chemical group that may be attached to a functional group and removed at a later stage to reveal the intact functional group. Examples of suitable protecting groups for various functional groups are described in T.W. Greene and P.G.M Wuts, Protective Groups in Organic Synthesis, 2d Ed., John Wiley and Sons (1991); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed. Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995).
• phrases "pharmaceutically acceptable salt(s)", as used herein, unless otherwise indicated, includes salts of acidic or basic groups which may be present in the compounds of the present invention.
• the compounds of the present invention that are basic in nature are capable of forming a wide variety of salts with various inorganic and organic acids.
• the acids that may be used to prepare pharmaceutically acceptable acid addition salts of such basic compounds of are those that form non-toxic acid addition salts, i.e ., salts containing pharmacologically acceptable anions, such as the hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, acid phosphate, isonicotinate, acetate, lactate, salicylate, citrate, acid citrate, tartrate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate and pamoate [ i.e ., 1,1'-methylene-bis-(2-hydroxy-3-naphtho
• Those compounds of the present invention that are acidic in nature are capable of forming base salts with various pharmacologically acceptable cations.
• Examples of such salts include the alkali metal or alkaline earth metal salts and, particularly, the calcium, magnesium, sodium and potassium salts of the compounds of the present invention.
• Certain compounds of the present invention may have asymmetric centers and therefore exist in different enantiomeric and diastereomic forms.
• This invention relates to the use of all optical isomers and stereoisomers of the compounds of the present invention, and mixtures thereof, and to all pharmaceutical compositions and methods of treatment that may employ or contain them.
• the present invention includes the compounds of the present invention, and the pharmaceutically acceptable salts thereof, wherein one or more hydrogen, carbon or other atoms are replaced by isotopes thereof.
• Such compounds may be useful as research and diagnostic tools in metabolism pharmacokinetic studies and in binding assays.
• the compounds of this invention including the compounds of formula I, are defined to include pharmaceutically acceptable derivatives or prodrugs thereof.
• a "pharmaceutically acceptable derivative or prodrug” means any pharmaceutically acceptable salt, ester, salt of an ester or other derivative of a compound of this invention that, upon administration to a recipient, is capable of providing (directly or indirectly) a compound of this invention or a metabolite or residue thereof.
• Particularly favored derivatives and prodrugs are those that increase the bioavailability of the compounds of this invention when such compounds are administered to a patient (e.g., by allowing an orally administered compound to be more readily absorbed into the blood), enhance delivery of the parent compound to a given biological compartment, increase solubility to allow administration by injection, alter metabolism or alter rate of excretion.
• Compounds of formula I can be converted into prodrugs through, for example, free amino, amido, hydroxy or carboxylic groups.
• prodrugs include compounds wherein an amino acid residue, or a polypeptide chain of two or more (e.g., two, three or four) amino acid residues is covalently joined through an amide or ester bond to a free amino, hydroxy or carboxylic acid group of a compound of formula I.
• amino acid residues include but are not limited to the 20 naturally occurring amino acids commonly designated by three letter symbols and also include 4-hydroxyproline, hydroxylysine, demosine, isodemosine, 3-methylhistidine, norvalin, beta-alanine, gamma-aminobutyric acid, citrulline homocysteine, homoserine, ornithine and methionine sulfone.
• prodrugs can be derivatized as amides or alkyl esters.
• the amide and ester moieties may incorporate groups including but not limited to ether, amine and carboxylic acid functionalities.
• Free hydroxy groups may be derivatized using groups including but not limited to hemisuccinates, phosphate esters, dimethylaminoacetates and phosphoryloxymethyloxycarbonyls, as outlined in D. Fleisher et al., Advanced Drug Delivery Reviews, vol. 19, p. 115 (1996).
• Carbamate prodrugs of hydroxy and amino groups are also included, as are carbonate prodrugs and sulfate esters of hydroxy groups.
• the compounds of this invention also include pharmaceutically acceptable salts of the compounds of formula I.
• pharmaceutically acceptable salt(s) includes salts of acidic or basic groups that may be present in the compounds of the present invention.
• the starting materials used in the present invention may require proper functional group protection before various modifications can take place, and deprotection after desired modifications are complete.
• Hydroxyl groups are generally protected as acetates or Cbz carbonates.
• the relative reactivity of various hydroxyl groups in the macrolide molecules of the general type claimed in this invention has been well established. Such differences in reactivity permit selective modification of different parts of the compounds of this invention.
• the synthetic process for preparation of compounds of formulae 5a-5c of the invention employs a 6-carbamoyl erythromycin A derivative 1 .
• Compounds of formula 1 and their syntheses are disclosed in US patent 4,826,820.
• the resulting compound is treated with DBU (1,8-diazabicyclo[5.4.0]undec-7-ene) and CDI (carbonyl diimidazole) to generate the 12-imidazolecarbonyloxy derivative 3.
• Treatment of this material with ammonia, hydrazine, or hydroxylamine generates compound 4a, 4b, or 4c, respectively. Removal of the protecting groups provides compounds 5a, 5b, or 5c.
• the compound of formula 1 is treated with 2 equivalents of acetic anhydride in the presence of a catalytic amount of 4-N,N,-dimethylaminopyridine in an inert solvent such as CH 2 Cl 2 at 0° to 50°C, preferably room temperature for 2 to 24 hours, preferably 12 hours (Scheme 1, Step 1).
• the diacetyl derivative of formula 2 is then treated with 1-2 eq of DBU and 1 eq of CDI in an inert solvent such as CH 2 Cl 2 at room temperature for 4 to 24 hours, preferably 12 hours (Scheme 1, Step 2) to obtain the 12-imidazolecarbonyloxy derivative of formula 3.
• the compound of formula 3 is the treated with 1-3 eq of ammonia, hydrazine or hydroxylamine in acetonitrile at 0° to 100°C, preferably 80° for 2-24 hours, preferably 12 hours (Scheme 1, Step 3) to obtain the compound of formula 4.
• the acetyl groups are removed from the compound of formula 4 by warming in methanol at room temperature to 80°C, preferably 50°C for 2-24 hours, preferably 12 hours (Scheme 1, Step 4) to obtain the compound of formula 5.
• Compounds of formula 1 containing an oxime at the 9 position can be obtained by converting a corresponding oxime lacking a carbamoyl at carbon 6, e.g., such as is described in US patent 4,990,602, to the corresponding cabamoyl 6 using methods known in the art, and to those described in US patent 4,826,820.
• compound 2 from Scheme 1 is treated with anhydrous HCI at 0°C to room temperature, preferably room temperature for 1 to 24 hours., preferably 12 hours, to prepare compound 3.
• the resulting compound is then treated with DMSO and EDCI (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) at 0°C to room temperature, preferably room temperature, for 1 to 24 hours, preferably 12 hours, to generate compound 8.
• oxidizing agents such as DMSO-oxalyl chloride or Dess-Martin periodinane reagent can also be employed.
• Compound 8 is treated with 1-2 equivalent of DBU in an inert solvent at 0 to 40°C, preferably 40°C, for 1 to 24 hours, preferably 12 hours.
• the resulting alcohol is then treated with NaH and carbonyl diimidazole (CDI) in THF at 0°C to room temperature, to generate compound 10.
• CDI carbonyl diimidazole
• Compound 10 is treated with R 2 -B-NH 2 in acetonitrile at room temperature to 80°C, preferably 80°C, for 1 to 24 hours, preferably 12 hours.
• compound 7 e.g., erythromycin A
• compound 14 is treated with ethylene carbonate and K 2 CO 3 in ethyleneglycol dimethyl ether at 50 to 85 °C, preferably 85°C, for 6 to 24 hours, preferably 24 hours, to give compound 14 .
• This is treated with acetic anhydride in THF at 0°C to room temperature, preferably 0°C, for 1 hr to 12 hours, preferably 12 hours to give compound 15.
• Compound 15 is then treated with trichloroacetyl isocyanate in dichloromethane at -10°C to room temperature, preferably 0°C, for 5 min to 5 hours, preferably 5 min.
• the resulting intermediates are treated with excess methanol to give compounds 16 and 17.
• Tricarbamate 17 is heated in aqueous acetonitrile at 50 to 80°C, preferably 80°C, for 3 to 24 hours, preferably 12 hours, to give compound 18.
• the cladinose sugar is cleaved as described above and the resulting alcohol is oxidized to ketone 19. This is then treated with excess base such as NaH and R-X 3 (X 3 is halogen, mesylate, tosylate, or other leaving group).
• the resulting intermediates are treated with MeOH to remove the C2' acetyl group to give compound 20 or 21.
• Dicarbamate 16 is treated with anhydrous HCI as described above to cleave the cladinose sugar and oxidized to give compound 22.
• the compounds of the present invention may have asymmetric carbon atoms and therefore exist in different enantiomeric and diastereomeric forms. Diastereomeric mixtures can be separated into their individual diastereomers on the basis of their physical chemical differences by methods known to those skilled in the art, for example, by chromatography or fractional crystallization. The use of all such isomers, including diastereomer mixtures and pure enantiomers, are considered to be part of the present invention.
• the compounds of the present invention that are basic in nature are capable of forming a wide variety of different salts with various inorganic and organic acids. Although such salts must be pharmaceutically acceptable for administration to mammals, it is often desirable in practice to initially isolate the compound of the present invention from the reaction mixture as a pharmaceutically unacceptable salt and then simply convert the latter back to the free base compound by treatment with an alkaline reagent and subsequently convert the latter free base to a pharmaceutically acceptable acid addition salt.
• the acid addition salts of the base compounds of this invention are readily prepared by treating the base compound with a substantially equivalent amount of the chosen mineral or organic acid in an aqueous solvent medium or in a suitable organic solvent, such as methanol or ethanol. Upon careful evaporation of the solvent, the desired solid salt is readily obtained.
• the desired salt can also be precipitated from a solution of the free base in an organic solvent by adding to the solution an appropriate mineral or organic acid.
• Those compounds of the present invention that are acidic in nature are capable of forming base salts with various cations.
• base salts include the alkali metal or alkaline-earth metal salts and particularly the sodium, amine and potassium salts. These salts are all prepared by conventional techniques.
• the chemical bases which are used as reagents to prepare the pharmaceutically acceptable base salts of this invention are those which form non-toxic base salts with the acidic compounds of the present invention.
• Such non-toxic base salts include those derived from such pharmacologically acceptable cations as sodium, potassium, calcium, magnesium, various amine cations, etc.
• These salts can easily be prepared by treating the corresponding acidic compounds with an aqueous solution containing the desired pharmacologically acceptable bases with cations such as sodium, potassium, calcium, magnesium, various amine cations, etc., and then evaporating the resulting solution to dryness, preferably under reduced pressure.
• they may also be prepared by mixing lower alkanolic solutions of the acidic compounds and the desired alkali metal alkoxide together, and then evaporating the resulting solution to dryness in the same manner as before.
• stoichiometric quantities of reagents are preferably employed in order to ensure completeness of reaction and maximum yields of the desired final product.
• the present invention includes all isotopically labelled forms of the compounds of formula I, and pharmaceutically acceptable salts and prodrugs thereof. Such isotopically labelled compounds are useful as research or diagnostic tools.
• the isotopically-labelled compounds and pharmaceutically acceptable salts thereof are identical to those of formula I but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature.
• Examples of isotopes that can be incorporated into compounds of this invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine and chlorine, such as 2 H, 3 H, 13 C, 14 C, 15 N, 18 O, 17 O, 35 S, 18 F and 36 Cl, respectively.
• isotopically labelled compounds of the present invention such as those into which radioactive isotopes such as 3 H and 14 C are incorporated, are useful in drug and/or substrate tissue distribution assays.
• Tritiated, i.e., 3 H, and carbon-14, i.e., 14 C, isotopes are particularly preferred for their ease of preparation and detectability.
• Substitution with heavier isotopes such as deuterium, i.e., 2 H can afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and hence may be preferred in some circumstances.
• Isotopically labelled compounds of formula I of this invention and prodrugs thereof can generally be prepared by carrying out the procedures disclosed in the Scheme(s) and/or in the Example(s) below and substituting a readily available isotopically labelled reagent for a non-isotopically labelled reagent.
• the antibacterial and antiprotozoa activity of the compounds of the present invention against bacterial and protozoa pathogens is demonstrated by the compound's ability to inhibit growth of defined strains of human (Assay I) or animal (Assays II and III) pathogens.
• Assay I employs conventional methodology and interpretation criteria and is designed to provide direction for chemical modifications that may lead to compounds that circumvent defined mechanisms of macrolide resistance.
• Assay I a panel of bacterial strains is assembled to include a variety of target pathogenic species, including representatives of macrolide resistance mechanisms that have been characterized. Use of this panel enables the chemical structure/activity relationship to be determined with respect to potency, spectrum of activity, and structural elements or modifications that may be necessary to obviate resistance mechanisms.
• Bacterial pathogens that comprise the screening panel are shown in the table below. In many cases, both the macrolide-susceptible parent strain and the macrolide-resistant strain derived from it are available to provide a more accurate assessment of the compound's ability to circumvent the resistance mechanism.
• Strains that contain the gene with the designation of ermA/ermB/ermC are resistant to macrolides, lincosamides, and streptogramin B antibiotics due to modifications (methylation) of 23S rRNA molecules by an Erm methylase, thereby generally prevent the binding of all three structural classes.
• Two types of macrolide efflux have been described; msrA encodes a component of an efflux system in staphylococci that prevents the entry of macrolides and streptogramins while mefA/E encodes a transmembrane protein that appears to efflux only macrolides.
• Inactivation of macrolide antibiotics can occur and can be mediated by either a phosphorylation of the 2'-hydroxyl ( mph ) or by cleavage of the macrocyclic lactone (esterase).
• the strains may be characterized using conventional polymerase chain reaction (PCR) technology and/or by sequencing the resistance determinant.
• PCR polymerase chain reaction
• the use of PCR technology in this application is described in J. Sutcliffe et al., "Detection Of Erythromycin-Resistant Determinants By PCR", Antimicrobial Agents and Chemotherapy, 40(11), 2562-2566 (1996).
• the assay is performed in microtiter trays and interpreted according to Performance Standards for Antimicrobial Disk Susceptibility Tests - Sixth Edition; Approved Standard, published by The National Committee for Clinical Laboratory Standards (NCCLS) guidelines; the minimum inhibitory concentration (MIC) is used to compare strains.
• Compounds are initially dissolved in dimethylsulfoxide (DMSO) as 40 mg/ml stock solutions.
• DMSO dimethylsulfoxide
• Assay II is utilized to test for activity against Pasteurella multocida and Assay III is utilized to test for activity against Pasteurella haemolytica.
• This assay is based on the liquid dilution method in microliter format.
• a single colony of P. multocida (strain 59A067) is inoculated into 5 ml of brain heart infusion (BHI) broth.
• the test compounds are prepared by solubilizing 1 mg of the compound in 125 ⁇ l of dimethylsulfoxide (DMSO). Dilutions of the test compound are prepared using uninoculated BHI broth. The concentrations of the test compound used range from 200 ⁇ g/ml to 0.098 ⁇ g/ml by two-fold serial dilutions.
• the P. multocida inoculated BHI is diluted with uninoculated BHI broth to make a 10 4 cell suspension per 200 ⁇ l.
• the BHI cell suspensions are mixed with respective serial dilutions of the test compound, and incubated at 37°C for 18 hours.
• the minimum inhibitory concentration (MIC) is equal to the concentration of the compound exhibiting 100% inhibition of growth of P . multocida as determined by comparison with an uninoculated control.
• This assay is based on the agar dilution method using a Steers Replicator. Two to five colonies isolated from an agar plate are inoculated into BHI broth and incubated overnight at 37°C with shaking (200 rpm). The next morning, 300 ⁇ l of the fully grown P. haemolytica preculture is inoculated into 3 ml of fresh BHI broth and is incubated at 37°C with shaking (200 rpm). The appropriate amounts of the test compounds are dissolved in ethanol and a series of two-fold serial dilutions are prepared. Two ml of the respective serial dilution is mixed with 18 ml of molten BHI agar and solidified. When the inoculated P .
• haemolytica culture reaches 0.5 McFarland standard density, about 5 ⁇ l of the P. haemolytica culture is inoculated onto BHI agar plates containing the various concentrations of the test compound using a Steers Replicator and incubated for 18 hours at 37°C. Initial concentrations of the test compound range from 100-200 ⁇ g/ml. The MIC is equal to the concentration of the test compound exhibiting 100% inhibition of growth of P. haemolytica as determined by comparison with an uninoculated control.
• the in vivo activity of the compounds of formula (I) can be determined by conventional animal protection studies well known to those skilled in the art, usually carried out in mice.
• mice are allotted to cages (10 per cage) upon their arrival, and allowed to acclimate for a minimum of 48 hours before being used.
• Animals are inoculated with 0.5 ml of a 3 x 10 3 CFU/ml bacterial suspension ( P . multocida strain 59A006) intraperitoneally.
• Each experiment has at least 3 non-medicated control groups including one infected with 0.1X challenge dose and two infected with 1X challenge dose; a 10X challenge data group may also be used.
• all mice in a given study can be challenged within 30-90 minutes, especially if a repeating syringe (such as a Cornwall® syringe) is used to administer the challenge.
• a repeating syringe such as a Cornwall® syringe
• the first compound treatment is given. It may be necessary for a second person to begin compound dosing if all of the animals have not been challenged at the end of 30 minutes.
• the routes of administration are subcutaneous or oral doses. Subcutaneous doses are administered into the loose skin in the back of the neck whereas oral doses are given by means of a feeding needle. In both cases, a volume of 0.2 ml is used per mouse. Compounds are administered 30 minutes, 4 hours, and 24 hours after challenge. A control compound of known efficacy administered by the same route is included in each test. Animals are observed daily, and the number of survivors in each group is recorded. The P. multocida model monitoring continues for 96 hours (four days) post challenge.
• the PD 50 is a calculated dose at which the compound tested protects 50% of a group of mice from mortality due to the bacterial infection which would be lethal in the absence of drug treatment.
• the compounds of formula 1, and the pharmaceutically acceptable salts, prodrugs, tautomers, and solvates thereof may be administered through oral, parenteral, topical, or rectal routes in the treatment of bacterial and protozoa infections.
• these compounds are most desirably administered in dosages ranging from about 0.2 mg per kg body weight per day (mg/kg/day) to about 200 mg/kg/day in single or divided doses (i.e., from 1 to 4 doses per day), although variations will necessarily occur depending upon the species, weight and condition of the subject being treated and the particular route of administration chosen.
• a dosage level that is in the range of about 4 mg/kg/day to about 50 mg/kg/day is most desirably employed. Variations may nevertheless occur depending upon the species of mammal, fish or bird being treated and its individual response to said medicament, as well as on the type of pharmaceutical formulation chosen and the time period and interval at which such administration is carried out. In some instances, dosage levels below the lower limit of the aforesaid range may be more than adequate, while in other cases still larger doses may be employed without causing any harmful side effects, provided that such larger doses are first divided into several small doses for administration throughout the day.
• the active compounds may be administered alone or in combination with pharmaceutically acceptable carriers or diluents by the routes previously indicated, and such administration may be carried out in single or multiple doses. More particularly, the active compounds may be administered in a wide variety of different dosage forms, i.e., they may be combined with various pharmaceutically acceptable inert carriers in the form of tablets, capsules, lozenges, troches, hard candies, powders, sprays, creams, salves, suppositories, jellies, gels, pastes, lotions, ointments, aqueous suspensions, injectable solutions, elixirs, syrups, and the like.
• Such carriers include solid diluents or fillers, sterile aqueous media and various non-toxic organic solvents, etc.
• oral pharmaceutical compositions can be suitably sweetened and/or flavored.
• the active compounds are present in such dosage forms at concentration levels ranging from about 5.0% to about 70% by weight.
• tablets containing various excipients such as microcrystalline cellulose, sodium citrate, calcium carbonate, dicalcium phosphate and glycine may be employed along with various disintegrants such as starch (and preferably corn, potato or tapioca starch), alginic acid and certain complex silicates, together with granulation binders like polyvinylpyrrolidone, sucrose, gelatin and acacia.
• disintegrants such as starch (and preferably corn, potato or tapioca starch), alginic acid and certain complex silicates, together with granulation binders like polyvinylpyrrolidone, sucrose, gelatin and acacia.
• lubricating agents such as magnesium stearate, sodium lauryl sulfate and talc are often very useful for tabletting purposes.
• compositions of a similar type may also be employed as fillers in gelatin capsules; preferred materials in this connection also include lactose or milk sugar as well as high molecular weight polyethylene glycols.
• the active compound may be combined with various sweetening or flavoring agents, coloring matter or dyes, and, if so desired, emulsifying and/or suspending agents as well, together with such diluents as water, ethanol, propylene glycol, glycerin and various like combinations thereof.
• solutions of an active compound in either sesame or peanut oil or in aqueous propylene glycol may be employed.
• the aqueous solutions should be suitably buffered (preferably pH greater than 8) if necessary and the liquid diluent first rendered isotonic.
• These aqueous solutions are suitable for intravenous injection purposes.
• the oily solutions are suitable for intraarticular, intramuscular and subcutaneous injection purposes. The preparation of all these solutions under sterile conditions is readily accomplished by standard pharmaceutical techniques will known to those skilled in the art.
• the active compounds may be administered in the feed of the animals or orally as a drench composition.
• the active compounds may also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles.
• Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines.
• the active compounds may also be coupled with soluble polymers as targetable drug carriers.
• soluble polymers can include polyvinylpyrrolidone, pyran copolymer, polyhydroxypropylmethacrylamide phenyl, polyhydroxyethylaspartamide-phenol, or polyethyleneoxide-polylysine substituted with palmitoylresidues.
• the active compounds may be coupled to a class of biodegradable polymers useful in achieving controlled release of a drug, for example, polylactic acid, polyglycolic acid, copolymers of polylactic and polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates and cross-linked or amphipathic block copolymers of hydrogels.
• a drug for example, polylactic acid, polyglycolic acid, copolymers of polylactic and polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates and cross-linked or amphipathic block copolymers of hydrogels.

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• 2001
  • 2001-04-03 EP EP01303172A patent/EP1146051A3/de not_active Withdrawn
  • 2001-04-04 JP JP2001105662A patent/JP2001348397A/ja active Pending
  • 2001-04-06 CA CA002343344A patent/CA2343344A1/en not_active Abandoned
  • 2001-04-09 MX MXPA01003647A patent/MXPA01003647A/es unknown
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• 2002
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